P635: TARGETING THE ONE-CARBON METABOLISM AS NOVEL THERAPEUTIC STRATEGY IN CHRONIC LYMPHOCYTIC LEUKEMIA
Authors: E. Viry, A. Largeot, S. Gonder, S. Chemlal, N. Kiweler, J. Meiser, T. Helleday, E. Moussay, J. Paggetti
Journal: HemaSphere (2022)

Background: Chronic Lymphocytic Leukemia (CLL), the most common type of leukemia in adults, is characterized by the clonal expansion of CD5+ CD19+ B cells. CLL progression is highly dependent on complex interactions with non-malignant cells of the tumor microenvironment. Despite great advance in the standard of care in the last decades, there are still unmet medical needs (long-life treatment, resistance…). Altered cellular metabolism has emerged as a hallmark of cancer by sustaining the uncontrolled growth of cancer cells. The one-carbon pathway is a major driver for tumor proliferation, providing building blocks for biosynthesis of nucleotides through pyrimidines (dTTP) and purines synthesis, redox metabolism (GSH), and energy balance by suppling ATP and NADPH to cells (A). Understanding the metabolic reprogramming occurring in cancer, notably in CLL, may provide insights to support the development of novel therapies. Aims: We aim to get insights into one-carbon metabolism regulation in CLL, identify new therapeutic targets and find new treatment options for CLL patients. Methods: Through a collaboration with a biotech company that develops inhibitors of the one-carbon metabolism, we tested new MTHFD1/2 inhibitors, which we believe, are promising in the context of CLL. The cytotoxic activity of the inhibitors was evaluated on a panel of murine and human CLL cell lines and primary cells, and also on others B cell malignancies such as Mantle Cell Lymphoma (MCL), Diffuse Large B-Cell Lymphoma (DLBCL), and Multiple Myeloma (MM). The molecular mechanisms sustaining the cytotoxic activity were evaluated by performing metabolic tracing, rescue experiments and CRISPR/Cas9 KO. Results: We showed a strong expression of both MTHFD1 and 2 in all the cancer cell lines tested and in stimulated primary human CLL cells. A higher expression of both enzymes was also observed in primary CLL cells, isolated from the Eµ-TCL1 murine model of CLL compared to normal B cells. In vitro treatment with the inhibitors efficiently reduced cell viability of CLL cell lines and primary cells, as for MCL and DLBCL cell lines, at low nanomolar dose while no effect was observed on MM cell lines (B). This effect is associated with a blockade of cell proliferation. Using 13C-serine isotope tracing, we could showed that the inhibitors significantly reduced ATP production from serine in CLL cells, probably resulting in altered nucleotides biosynthesis. By performing rescue experiments with thymidine or hypoxanthine, we confirmed that the cytotoxic effect of the inhibitors on CLL cells is mediated through a defect in pyrimidine synthesis (C). Interestingly, we demonstrated that the resistant MM cell lines overexpressed the SHMT1 enzyme, and could be sensitized to MTHFD1/2 inhibition by combination therapy with SHMT1/2 inhibitor (D). Taken together these results suggest that SHMT1 may be a key player in the response to MTHFD1/2 inhibition. Furthermore, we have previously reported that CLL induces a dramatic remodeling of tumor microenvironment, notably leading to the accumulation of highly immunosuppressive Tregs (Wierz et al., Blood, 2018). We showed that both enzymes are overexpressed in those highly immunosuppressive Tregs, and that MTHD1/2 inhibitors are able to reduce the viability of ex vivo polarized Tregs. Image:Summary/Conclusion: MTHFD1/2 inhibitors display a high cytotoxic activity in CLL and other B-cell malignancies by impairing cell proliferation through a defect in pyrimidine synthesis. These inhibitors also exhibit cytotoxic effect on activated Treg cells ex vivo, reinforcing its therapeutic potential for CLL treatment.